Differential regulation of endothelium behavior by progesterone and medroxyprogesterone acetate

Medroxyprogesterone acetate (MPA) is a synthetic progestin commonly used in hormone replacement therapy (HRT). The aim of this research was to study and compare the effect of progesterone (Pg) and MPA on the regulation of cellular events associated with vascular homeostasis and disease. Platelet adhesion to endothelial cells (ECs), nitric oxide (NO) production, and cell migration were studied using murine ECs in vitro exposed to the progestins. After 7 min of treatment, MPA significantly inhibited NO synthesis with respect to control values; meanwhile, Pg markedly increased vasoactive production. In senile ECs, the stimulatory action of Pg decreases; meanwhile, MPA maintained its ability to inhibit NO synthesis. The presence of RU486 antagonized the action of each steroid. When ECs were preincubated with PD98059 (MAPK inhibitor) or chelerythrine (protein kinase C (PKC) inhibitor) before Pg or MPA treatment, the former totally suppressed the steroid action, but the PKC antagonist did not affect NO production. In the presence of a PI3K inhibitor (LY294002), a partial reduction in Pg effect and a reversal of MPA action were detected. Using indomethacin, the contribution of the cyclooxygenase (COX) pathway was also detected. On platelet adhesion assays, Pg inhibited and MPA stimulated platelet adhesion to ECs. Under inflammatory conditions, Pg prevented platelet adhesion induced by lipopolysaccharide (LPS); meanwhile, MPA potentiated the stimulatory action of LPS. Finally, although both steroids enhanced migration of ECs, MPA exhibited a greater effect. In conclusion, the data presented in this research provide evidence of a differential regulation of vascular function by Pg and MPA.

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Vascular responses to sodium nitroprusside in the human fetal-placental circulation

Reproduction Fertility and Development ◽

10.1071/rd9951557 ◽

1995 ◽

Vol 7 (6) ◽

pp. 1557 ◽

Cited By ~ 6

Author(s):

MA Read ◽

WB Giles ◽

IM Leitch ◽

AL Boura ◽

WA Walters

Keyword(s):

Sodium Nitroprusside ◽

Vascular Function ◽

No Production ◽

Low Oxygen ◽

Experimental Conditions ◽

Arterial Perfusion ◽

Basal Tone ◽

Intra Uterine Growth Retardation ◽

Synthase Inhibitor

This study examined the activity of sodium nitroprusside (SNP) in the human fetal-placental circulation in vitro in pathological and experimental conditions in which vascular function may be impaired. SNP (13-3400 nM) caused a concentration-dependent reduction in fetal arterial perfusion pressure (FAP) in Krebs' perfused placental cotyledons, at basal tone and following pre-constriction with prostaglandin F2 alpha (PGF2 alpha). SNP-induced reduction in FAP in the PGF2 alpha pre-constricted fetal-placental circulation was enhanced approximately six-fold (5.85) in those placentae pre-treated with the nitric oxide (NO) synthase inhibitor N omega-nitro-L-arginine (100 microM). Reductions in FAP in the preconstricted fetal-placental vasculature caused by SNP were not altered by prior infusion of ouabain (100 nM) into the fetal circulation or during low oxygen perfusion (O2 tension < 50 mmHg). No differences were observed in the responses obtained to SNP in placentae obtained from women with normotensive pregnancies or those associated with (i) pregnancy-induced hypertension, (ii) intra-uterine growth retardation, or (iii) an elevated umbilical-artery Doppler-ultrasound systolic/diastolic ratio, in either preconstricted placentae or those at basal tone. These findings are consistent with an up-regulation of guanylate cyclase/cGMP-mediated vasodilatation in the fetal-placental vasculature following complete blockade of endogenous NO production.

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Abstract P275: Deletion Of Endothelial Leptin Receptor Elevates Blood Pressure And Impairs Endothelial-dependent Relaxation Via Upregulation Of Endothelial Glycolytic Enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3)

Hypertension ◽

10.1161/hyp.78.suppl_1.p275 ◽

2021 ◽

Vol 78 (Suppl_1) ◽

Author(s):

Reem T. Atawia ◽

Jessica L. Faulkner ◽

Simone Kennard ◽

Vinay Mehta ◽

Galina Antonova ◽

...

Keyword(s):

Blood Pressure ◽

Vascular Function ◽

Leptin Receptor ◽

Ex Vivo ◽

Glycolytic Enzyme ◽

No Production ◽

Cell Type ◽

No Bioavailability ◽

Endothelial Homeostasis

The adipokine leptin plays a crucial role in blood pressure (BP) regulation notably by exerting pressor effects centrally via sympatho-activation and depressor effects via direct activation of its receptor (LepR) peripherally resulting in nitric oxide (NO)-mediated vasodilation. However, the predominant effects and cell type responsible for leptin-mediated NO production is not clearly understood. Herein, we examined the effect of selective deletion of LepR in endothelial cells (LepR EC-/- , KO) on BP and vascular function. BP recording via radiotelemetry in male KO and WT (LepR EC+/+ ) mice revealed significant increases in diastolic and mean arterial pressure in KO mice (DBP, WT: 90.2±2.1 vs. KO: 100.1±3.6; MAP, WT: 105.7±2.1 vs. KO: 113.7±2.6 mmHg, n=5, p<0.05). There was no difference in Systolic blood pressure or heart rate between KO and WT. Leptin infusion (0.9mg/kg/day,7 days) elicits a significant increase in BP of WT but not KO mice (DBP, WT: 89.2± 2.6 vs WT+Leptin 95.7±3.3; MAP, WT: 104 ±2.8 vs WT+Leptin: 110 ±2.7, n=5, p<0.05). We quantified sympathetic contribution to BP elevation by measuring BP response to glanglionic blockade (Hexamethonium). At baseline, KO mice exhibited a lower BP response than WT supporting a reduced neurogenic control of BP regulation in KO mice. Vascular contribution to high BP was investigated using wire myography in thoracic aorta. LepR deficiency impaired endothelial-dependent relaxation (EDR) to acetylcholine (n=7, p<0.05). L-NAME completely abolished EDR in KO and WT indicating that EC LepR deficiency reduced NO bioavailability. Recent evidence presents PFKFB3-mediated EC glycolysis as a new regulator of endothelial homeostasis. We found that aortic EC from KO exhibited increased PFKFB3 mRNA expression (p=0.065) and PFKFB3 inhibition restored EDR in KO. Remarkably, overexpression of PFKFB3 increased EC glycolysis in vitro and impaired EDR in WT aortic rings ex vivo . Collectively, our data suggest that impaired endothelial leptin receptor signaling induces a PFKFB3-dependent hyper-glycolytic phenotype resulting in NO deficiency and endothelial dysfunction that predisposes to higher BP regardless the reduced sympatho-activation which might prevent the increase in BP induced by exogenous leptin.

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High Glucose-enhanced Acetylcholine Stimulated CGMP Masks Impaired Vascular Reactivity in Tail Arteries from Short-Term Hyperglycemic Rats

International Journal of Experimental Diabetes Research ◽

10.1155/edr.2000.69 ◽

2000 ◽

Vol 1 (1) ◽

pp. 69-79 ◽

Cited By ~ 5

Author(s):

Marwan Hamaty ◽

Cristina B. Guzmán ◽

Mary F. Walsh ◽

Ann M. Bode ◽

Joseph Levy ◽

...

Keyword(s):

Oxidative Stress ◽

High Glucose ◽

Vascular Function ◽

No Production ◽

Short Term ◽

Contractile Responses ◽

Vascular Responses ◽

Short Term Exposure ◽

Endothelium Dependent Relaxation

Impaired vascular endothelium-dependent relaxation and augmented contractile responses have been reported in several models of long-term hyperglycemia. However, the effects of short-term ambient hyperglycemia are poorly understood. Since oxidative stress has been implicated as a contributor to impaired vascular function, we investigated the following:Aims: (1) the effects of high glucose exposurein vitro(7 – 10 days) on vascular relaxation to acetylcholine (Ach) and contractility to norepinephrine (NE) and KCl; (2) if NO-dependent cGMP generation is affected under these conditions; and (3) aortic redox status.Methods: Non-diabetic rat tail artery rings were incubated in normal (5mM) (control NG) or high (20mM) glucose buffer (control HG). Vascular responses to Ach, NE and KCl were compared to those of streptozotocin (SZ) diabetic animals in the same buffers (diabetic NG, diabetic HG). Ach stimulated cGMP levels were quantitated as an indirect assessment of endothelial nitric oxide (NO) production and oxidative stress evaluated by measuring vascular glutathione and oxidized glutathione.Results: Rings from diabetic rats in NG showed impaired relaxation to Ach (P= 0.002) but relaxed normally, when maintained in HG. Similarly, contractile responses to NE were attenuated in diabetic rings in NG but similar to controls in HG. HG markedly augmented maximal contraction to KCl compared to control and diabetic vessels in NG (P< 0.0001). Diabetic vessels in a hyperosmolar, but normoglycemic, milieu respond like those in HG.in vitro, HG for 2 hours changed neither relaxation nor contractile responses to NE and KCl in control rings. Basal cGMP levels were lower in aortae from diabetic animals pre-incubated in NG than in HG/LG or in control rings in NG (P< 0.05). cGMP responses to Ach were exaggerated in diabetic vessels in HG (P= 0.035vs. control NG,P= 0.043vs. diabetic NG) but not different between control and diabetic rings in NG. Vessels from diabetic animals had lower levels of GISH (P< 0.0001) and higher levels of GSSG (P< 0.0001) indicating oxidative stress.Conclusions: Our data indicate that endothelium dependent relaxation is altered early in the diabetic state and that increased NO responses may compensate for augmented oxidative stress but the lack of effect of short-term exposure of normal vessels to HG suggests that short-term hyperglycemiaper sedoes not cause abnormal vascular responses.

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Statins Reduce Lipopolysaccharide-Induced Cytokine and Inflammatory Mediator Release in an In Vitro Model of Microglial-Like Cells

Mediators of Inflammation ◽

10.1155/2017/2582745 ◽

2017 ◽

Vol 2017 ◽

pp. 1-10 ◽

Cited By ~ 12

Author(s):

A. J. McFarland ◽

A. K. Davey ◽

S. Anoopkumar-Dukie

Keyword(s):

In Vitro Model ◽

No Production ◽

In Vivo Models ◽

Cellular Models ◽

Reductase Inhibitors ◽

Inflammatory Conditions ◽

Vitro Model ◽

Coa Reductase

The anti-inflammatory effects of statins (HMG-CoA reductase inhibitors) within the cardiovascular system are well-established; however, their neuroinflammatory potential is unclear. It is currently unknown whether statins’ neurological effects are lipid-dependent or due to pleiotropic mechanisms. Therefore, the assumption that all statin compounds will have the same effect within the central nervous system is potentially inappropriate, with no studies to date having compared all statins in a single model. Thus, the aim of this study was to compare the effects of the six statins (atorvastatin, fluvastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin) within a single in vitro model of neuroinflammation. To achieve this, PMA-differentiated THP-1 cells were used as surrogate microglial cells, and LPS was used to induce inflammatory conditions. Here, we show that pretreatment with all statins was able to significantly reduce LPS-induced interleukin (IL)-1βand tumour necrosis factor (TNF)-αrelease, as well as decrease LPS-induced prostaglandin E2 (PGE2). Similarly, global reactive oxygen species (ROS) and nitric oxide (NO) production were decreased following pretreatment with all statins. Based on these findings, it is suggested that more complex cellular models should be considered to further compare individual statin compounds, including translation into in vivo models of acute and/or chronic neuroinflammation.

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Effects of Bupranolol, a New β-Blocker, on Platelet Functions of Rabbit and Human in Vitro

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648052 ◽

1976 ◽

Vol 36 (02) ◽

pp. 376-387 ◽

Cited By ~ 3

Author(s):

Teruhiko Umetsu ◽

Kazuko Sanai ◽

Tadakatsu Kato

Keyword(s):

Platelet Aggregation ◽

Platelet Adhesion ◽

Human Platelets ◽

Rabbit Platelet ◽

Rabbit Platelets ◽

Β Blocker ◽

Platelet Aggregates ◽

Induced Aggregation ◽

Platelet Functions

SummaryThe effects of bupranolol, a new β-blocker, on platelet functions were investigated in vitro in rabbits and humans as compared with propranolol, a well-known β-blocker. At first, the effect of adrenaline on ADP-induced rabbit platelet aggregation was studied because adrenaline alone induces little or no aggregation of rabbit platelets. Enhancement of ADP-induced rabbit platelet aggregation by adrenaline was confirmed, as previously reported by Sinakos and Caen (1967). In addition the degree of the enhancement was proved to be markedly affected by the concentration of ADP and to increase with decreasing concentration of ADP, although the maximum aggregation (percent) was decreased.Bupranolol and propranolol inhibited the (adrenaline-ADP-)induced aggregation of rabbit platelets, bupranolol being approximately 2.4–3.2 times as effective as propranolol. Bupranolol stimulated the disaggregation of platelet aggregates induced by a combination of adrenaline and ADP, but propranolol did not. Platelet adhesion in rabbit was also inhibited by the β-blockers and bupranolol was more active than propranolol. With human platelets, aggregation induced by adrenaline was inhibited by bupranolol about 2.8–3.3 times as effectively as propranolol.From these findings. We would suggest that bupranolol might be useful for prevention or treatment of thrombosis.

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In Vitro Assay of Platelet Adhesiveness with Washed and Tanned Human Red Cells

Thrombosis and Haemostasis ◽

10.1055/s-0038-1651281 ◽

1968 ◽

Vol 20 (03/04) ◽

pp. 384-396 ◽

Cited By ~ 1

Author(s):

G Zbinden ◽

S Tomlin

Keyword(s):

Platelet Adhesion ◽

Sodium Citrate ◽

Blood Platelets ◽

In Vitro Assay ◽

Red Cells ◽

Platelet Adhesiveness ◽

Vitro Assay ◽

In Vitro System ◽

Human Red Cells

SummaryAn in vitro system is described in which adhesion of blood platelets to washed and tannic acid-treated red cells was assayed quantitatively by microscopic observation. ADP, epinephrine and TAME produced a reversible increase in platelet adhesiveness which was antagonized by AMP. With Evans blue, polyanetholsulfonate, phthalanilide NSC 38280, thrombin and heparin at concentrations above 1-4 u/ml the increase was irreversible. The ADP-induced increase in adhesiveness was inhibited by sodium citrate, EDTA, AMP, ATP and N-ethylmaleimide. EDTA, AMP and the SH-blocker N-ethylmaleimide also reduced spontaneous platelet adhesion to red cells. No significant effects were observed with adenosine, phenprocoumon, 5-HT, phthalanilide NSC 57155, various estrogens, progestogens and fatty acids, acetylsalicylic acid and similarly acting agents, hydroxylamine, glucose and KCN. The method may be useful for the screening of thrombogenic and antithrombotic properties of drugs.

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Perturbation of Platelet Adhesion to Endothelial Cells by Plasminogen Activation In Vitro

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657655 ◽

1997 ◽

Vol 78 (02) ◽

pp. 934-938 ◽

Cited By ~ 5

Author(s):

Hsiun-ing Chen ◽

Yueh-I Wu ◽

Yu-Lun Hsieh ◽

Guey-Yueh Shi ◽

Meei-Jyh Jiang ◽

...

Keyword(s):

Extracellular Matrix ◽

Endothelial Cells ◽

Platelet Adhesion ◽

Treatment Time ◽

Shape Change ◽

Umbilical Vein ◽

Tissue Type ◽

Apical Surface ◽

Plasminogen Activation

SummaryTo investigate whether the endothelium-platelet interactions may be altered by plasminogen activation, cultured human umbilical vein endothelial cells (ECs) were treated with tissue-type plasminogen activator (t-PA) in the presence of plasminogen, and platelet adhesion to ECs was subsequently measured by using a tapered flow chamber. Our results demonstrated that platelets adhered more readily to t-PA treated EC monolayer than to the control monolayer at all shear stress levels tested. This phenomenon was treatment time-dependent and dose-dependent, and it could be blocked by adding plasmin inhibitors, such as e-amino caproic acid and aprotinin. Adherent platelets on t-PA treated EC monolayer underwent more severe shape change than those on the control monolayer. While the extracellular matrix directly treated with t-PA attracted less platelets than the control matrix did, platelet adhesion to the matrix that was produced by t-PA-treated ECs was unaltered. These data suggest that t-PA treatment on ECs compromised antiplatelet-adhesion capability on their apical surface without altering the reactivity of their extracellular matrix towards platelets.

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Desmopressin (DDAVP) Enhances Platelet Adhesion to the Extracellular Matrix of Cultured Human Endothelial Cells through Increased Expression of Tissue Factor

Thrombosis and Haemostasis ◽

10.1055/s-0038-1656088 ◽

1997 ◽

Vol 77 (05) ◽

pp. 0975-0980 ◽

Cited By ~ 23

Author(s):

Angel Gálvez ◽

Goretti Gómez-Ortiz ◽

Maribel Díaz-Ricart ◽

Ginés Escolar ◽

Rogelio González-Sarmiento ◽

...

Keyword(s):

Extracellular Matrix ◽

Endothelial Cells ◽

Tissue Factor ◽

Platelet Adhesion ◽

Umbilical Vein ◽

Procoagulant Activity ◽

Experimental Conditions ◽

Chromogenic Assay

SummaryThe effect of desmopressin (DDAVP) on thrombogenicity, expression of tissue factor and procoagulant activity (PCA) of extracellular matrix (ECM) generated by human umbilical vein endothelial cells cultures (HUVEC), was studied under different experimental conditions. HUVEC were incubated with DDAVP (1, 5 and 30 ng/ml) and then detached from their ECM. The reactivity towards platelets of this ECM was tested in a perfusion system. Coverslips covered with DD A VP-treated ECMs were inserted in a parallel-plate chamber and exposed to normal blood anticoagulated with low molecular weight heparin (Fragmin®, 20 U/ml). Perfusions were run for 5 min at a shear rate of 800 s1. Deposition of platelets on ECMs was significantly increased with respect to control ECMs when DDAVP was used at 5 and 30 ng/ml (p <0.05 and p <0.01 respectively). The increase in platelet deposition was prevented by incubation of ECMs with an antibody against human tissue factor prior to perfusion. Immunofluorescence studies positively detected tissue factor antigen on DDAVP derived ECMs. A chromogenic assay performed under standardized conditions revealed a statistically significant increase in the procoagulant activity of the ECMs produced by ECs incubated with 30 ng/ml DDAVP (p <0.01 vs. control samples). Northern blot analysis revealed increased levels of tissue factor mRNA in extracts from ECs exposed to DDAVP. Our data indicate that DDAVP in vitro enhances platelet adhesion to the ECMs through increased expression of tissue factor. A similar increase in the expression of tissue factor might contribute to the in vivo hemostatic effect of DDAVP.

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Sex-dependent dysregulation of human neutrophil responses by bisphenol A

Environmental Health ◽

10.1186/s12940-020-00686-8 ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Wioletta Ratajczak-Wrona ◽

Marzena Garley ◽

Malgorzata Rusak ◽

Karolina Nowak ◽

Jan Czerniecki ◽

...

Keyword(s):

Nadph Oxidase ◽

Bisphenol A ◽

Total Concentration ◽

Nicotinamide Adenine Dinucleotide Phosphate ◽

Human Neutrophils ◽

Boyden Chamber ◽

No Production ◽

Pathogen Elimination ◽

Latex Beads

Abstract Background In the present study, we aimed to investigate selected functions of human neutrophils exposed to bisphenol A (BPA) under in vitro conditions. As BPA is classified among xenoestrogens, we compared its action and effects with those of 17β-estradiol (E2). Methods Chemotaxis of neutrophils was examined using the Boyden chamber. Their phagocytosis and nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) oxidase activity were assessed via Park’s method with latex beads and Park’s test with nitroblue tetrazolium. To assess the total concentration of nitric oxide (NO), the Griess reaction was utilized. Flow cytometry was used to assess the expression of cluster of differentiation (CD) antigens. The formation of neutrophil extracellular traps (NETs) was analyzed using a microscope (IN Cell Analyzer 2200 system). Expression of the investigated proteins was determined using Western blot. Results The analysis of results obtained for both sexes demonstrated that after exposure to BPA, the chemotactic capacity of neutrophils was reduced. In the presence of BPA, the phagocytic activity was found to be elevated in the cells obtained from women and reduced in the cells from men. Following exposure to BPA, the percentage of neutrophils with CD14 and CD284 (TLR4) expression, as well as the percentage of cells forming NETs, was increased in the cells from both sexes. The stimulatory role of BPA and E2 in the activation of NADPH oxidase was observed only in female cells. On the other hand, no influence of E2 on the expression of CD14 and CD284, chemotaxis, phagocytosis, and the amount of NET-positive neutrophils was found for both sexes. The study further showed that BPA intensified NO production and iNOS expression in the cells of both sexes. In addition, intensified expression of all tested PI3K-Akt pathway proteins was observed in male neutrophils. Conclusions The study demonstrated the influence of BPA on neutrophil functions associated with locomotion and pathogen elimination, which in turn may disturb the immune response of these cells in both women and men. Analysis of the obtained data showed that the effect of this xenoestrogen on the human neutrophils was more pronounced than E2.

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Interleukin 8 Elicits Rapid Physiological Changes in Neutrophils That Are Altered by Inflammatory Conditions

Journal of Innate Immunity ◽

10.1159/000514885 ◽

2021 ◽

pp. 1-17

Author(s):

Stefan Bernhard ◽

Stefan Hug ◽

Alexander Elias Paul Stratmann ◽

Maike Erber ◽

Laura Vidoni ◽

...

Keyword(s):

Systemic Inflammation ◽

Severe Trauma ◽

Induced Response ◽

Neutrophil Granulocytes ◽

Inflammatory Conditions ◽

Flow Cytometric ◽

Time Flow ◽

Flow Cytometric Measurement ◽

Detailed Kinetics

A sufficient response of neutrophil granulocytes stimulated by interleukin (IL)-8 is vital during systemic inflammation, for example, in sepsis or severe trauma. Moreover, IL-8 is clinically used as biomarker of inflammatory processes. However, the effects of IL-8 on cellular key regulators of neutrophil properties such as the intracellular pH (pH<sub>i</sub>) in dependence of ion transport proteins and during inflammation remain to be elucidated. Therefore, we investigated in detail the fundamental changes in pH<sub>i</sub>, cellular shape, and chemotactic activity elicited by IL-8. Using flow cytometric methods, we determined that the IL-8-induced cellular activity was largely dependent on specific ion channels and transporters, such as the sodium-proton exchanger 1 (NHE1) and non-NHE1-dependent sodium flux. Exposing neutrophils in vitro to a proinflammatory micromilieu with N-formyl-Met-Leu-Phe, LPS, or IL-8 resulted in a diminished response regarding the increase in cellular size and pH. The detailed kinetics of the reduced reactivity of the neutrophil granulocytes could be illustrated in a near-real-time flow cytometric measurement. Last, the LPS-mediated impairment of the IL-8-induced response in neutrophils was confirmed in a translational, animal-free human whole blood model. Overall, we provide novel mechanistic insights for the interaction of IL-8 with neutrophil granulocytes and report in detail about its alteration during systemic inflammation.

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